lncRNA XIST通过调控miR-337-3p/HOXC8 轴促进胃癌的发展进程

[摘要] 目的：探讨lncRNA XIST（XIST）通过调控miR-337-3p/HOXC8 分子轴介导胃癌发展进程的分子机制。方法：选取2013 年3 月至2018 年1 月河北省唐山市开滦总医院普通外科手术切除的58 例胃癌组织和对应的癌旁组织标本,以及人胃癌细胞系AGS、MGC803、HGC27 和人胃黏膜细胞GES-1,用qPCR检测胃癌组织和细胞系中XIST 和miR-337-3p 的表达水平。将XIST敲降载体、miR-337-3p mimics/inhibitor 和HOXC8 过表达载体转染进AGS细胞,用CCK-8、Transwell 实验检测AGS细胞的增殖及侵袭能力,用WB检测HOXC8 及上皮钙黏蛋白（E-cadherin）、神经钙黏蛋白（N-cadherin）和波形蛋白（vimentin）的表达水平。用双荧光素酶报告基因验证XIST、miR-337-3p 和HOXC8 的靶向关系。结果：XIST在胃癌组织和细胞系中高表达（均P<0.01）。敲降XIST 显著抑制AGS细胞的增殖、侵袭和EMT（P<0.05 或P<0.01）。XIST 靶向作用miR-337-3p 并下调其表达,HOXC8 是miR-337-3p 的靶基因。敲降XIST 通过靶向上调miR-337-3p 对HOXC8 表达的抑制作用,从而抑制AGS细胞增殖、侵袭和EMT（P<0.05 或P<0.01）。结论：敲降XIST 可抑制AGS 细胞增殖、侵袭和EMT,其作用机制是通过靶向miR-337-3p 并下调HOXC8的表达。     

血管生成和p16、p53蛋白表达与非小细胞肺癌淋巴结转移的相关性研究

目的　分析非小细胞肺癌 (NSCLC)淋巴结转移与血管生成及p16、p5 3蛋白表达的关系。方法　采用免疫组织化学方法对 187例NSCLC(74例腺癌、76例鳞癌和 37例腺鳞混合癌 )进行血管染色计数和p16、p5 3蛋白测定 ,并作相关临床因素分析。结果　非小细胞肺癌组织中微血管密度平均为 6 .8± 3.3,p16和p5 3蛋白阳性表达率分别为 4 7.6 %和 34.2 %。各组织类型非小细胞肺癌中淋巴结转移病例 (T2 N1～ 2 M0 )的微血管密度显著高于未转移者 (T2 N0 M0 ) ;在肺鳞癌和腺鳞癌中 ,p16和p5 3蛋白同时表达异常 (p16 -p5 3 )与微血管密度增高有关 ,但与淋巴结转移无关 ;而在肺腺癌中 ,p16和p5 3蛋白同时表达异常与淋巴结转移有关 ,但与微血管密度无关。微血管密度和p16、p5 3蛋白表达异常与年龄、性别、病理类型及分化程度均无明显相关。结论　各组织类型非小细胞肺癌的淋巴结转移均与活跃的血管生成密切相关 ,其中在肺鳞癌和腺鳞癌中 ,血管生成与p16和p5 3基因表达异常有关。而在肺腺癌中 ,血管生成和p16、p5 3蛋白异常表达是影响其淋巴结转移的两项独立的因素。     

p120-catenin亚型的表达减弱与肺癌的淋巴结转移明显相关

背景与目的 已有的研究表明,p120ctn(p120 catenin)在细胞粘附中起重要作用,与肿瘤发生关系密切.本研究探讨p120ctn哑型在肺鳞癌、腺癌中的表达及其与各临床病理因素的关系.方法应用RT-PCR方法检测肺鳞癌、腺痛及对应癌旁肺组织中p120ctn各亚型的表达情况,同时应用免疫荧光、Western-blot和RT-PCR方法检测了p120ctn各业型在肺癌PG细胞系BE1和LH7中的表达.结果 p120ctn的总mRNA及其亚型1和亚型3的mRNA在癌旁肺组织中的表达量要明显高于肺鳞、腺痛组织中的表达量(P<0.001,P<0.001,P=0.001).且亚型1 mRNA的表达缺失与淋巴结转移呈负相关(P=0.01),而亚型3 mRNA的表达缺失与癌组织的淋巴结转移呈正相关(P=0.029).高转移能力的BE1细胞中p120ctn亚型1和亚型3的表达减弱比LH7更为明显.结论 p120ctn亚型1和3 mRNA的表达减弱或缺失是肺癌中普遍存在的现象,且这种现象与淋巴结的转移有密切关系.     

胃癌转移相关miRNA的差异表达谱及miR-218的生物学分析

目的 探讨胃癌转移相关微小RNA（miRNA）的差异表达情况并进行miR-218的生物学分析。方法 采用实时荧光定量PCR（qPCR）及miRNA芯片法检测低转移潜能的胃癌细胞亚系（SGC7901-NM、MKN28-NM）与高转移潜能的胃癌细胞亚系（SGC7901-M、MKN28-M）间miRNA的差异表达。提取不同转移潜能胃癌细胞系和10例胃癌冰冻组织及相应的转移淋巴结中的总RNA,利用qPCR检测miR-218在不同细胞及组织中的表达情况。结果 对不同转移潜能的胃癌细胞亚系进行芯片检测发现,与SGC7901-NM细胞比较,SGC7901-M 细胞有47个分子表达下调,15个分子表达上调。与MKN28-NM细胞比较,MKN28-M细胞有41个分子表达下调,83个分子表达上调。在SGC7901-M及MKN28-M细胞中,34个分子表达均出现下降,11个分子表达均出现上升。对不同转移潜能的胃癌细胞亚系以及人永生化正常胃黏膜细胞系GES进行检测可以发现,4种不同转移潜能的胃癌细胞亚系中miR-218的表达均低于正常胃黏膜细胞系GES,差异有统计学意义（P＜0.05）,且在高转移潜能胃癌细胞亚系中miR-218的表达均低于低转移潜能胃癌细胞系,差异有统计学意义（P＜0.05）。胃癌转移淋巴结中miR-218的表达水平为0.23±0.02,低于胃癌原发灶的1.09±0.05,差异有统计学意义（P＜0.05）。结论 胃癌转移相关miRNA会出现差异表达情况,高转移潜能胃癌细胞中的miR-218表达水平上调可能与胃癌转移存在一定关系。     

以hsa-miR-491-3p为靶标的反义寡核苷酸对NCI-H446细胞生物学特性的影响

背景与目的:miRNAs是一类小分子RNA,参与转录后调控.前期研究结果显示,hsa-miR-491-3p在肺痛和肾痛组织中高表达,目前鲜见其功能研究的报道.本研宄旨在探讨以hsa-miR-491-3p为靶标的反义寡核苷酸对人小细胞肺癌NCI-H446细胞生物学特性的影响.方法:合成以hsa-miR-491-3p为靶标的反义寡核苷酸和随机对照序列,转染NCI-H446细胞.实验分4组:空白对照组(组1)、空白转染组(组2) ⑺婊?对照组(组3)和反义寡核苷酸组(组4).用四甲基偶氮唑蓝(MTT)法筛选最佳作用浓度,Hoechst33258染色检测细胞核的形态变化,流式细胞术检测细胞凋亡和周期变化,划痕实验检测细胞的迁移能力.结果:转染反义寡核苷酸后细胞生长受到抑制,最佳作用浓度为0.5 μmol/L.Hoechst33258染色各组的凋亡率分别为(6.67±0.58)%、(7.00±1.00)%、(6.67±1.53)%和(26.33±1.53)%,组4的凋亡率明显高于前3组,差异有统计学意义(P<0.001).流式细胞仪检测各组的凋亡率分别为(2.44±0.60)%、(2.59±0.85)%、(2.62±1.11)%和(5.22±0.40)%,组4的凋亡率明显高于前3组,差异有统计学意义(P<0.01),且G1期细胞增多,S期细胞减少.划痕实验显示各组细胞24 h的迁移距离分别为(196.88±39.13)、(163.06±21.28)、(225.49±23.42)和(77.41±6.07)μ m,组4细胞的迁移距离小于前3组(P<0.05),迁移能力减弱.结论:以hsa-miR-491-3p为靶标的反义寡核苷酸能够抑制NCI-H446细胞增殖,诱导其凋亡,并降低其侵袭能力.     

Correlation between survivin mRNA expression and lymph node metastasis in gastric cancer

Background Correlations between the malignant potential and prognosis of cancer and abnormal control mechanisms of apoptosis have been discovered in a variety of cancers. Survivin is a member of the inhibiting apoptosis protein family that is abundant in embryonic and carcinoma tissues. We measured the expression of survivin mRNA in gastric cancer to determine whether levels of survivin mRNA expression could serve as an index of malignancy.Methods Expression of survivin mRNA was measured in samples of both gastric cancer and noncancerous tissue from 107 patients. Survivin mRNA was detected by the real-time polymerase chain reaction (PCR) method, and then the relationship between the survivin mRNA level and histological diagnosis was analyzed.Results Expression of survivin mRNA was observed in 105 of 107 cancerous tissues and in 101 of 107 noncancerous tissues. The Mean value of survivin mRNA expression in cancerous tissue was 5.18 ± 1.30, significantly higher (P 0.01) than that in noncancerous tissue, at 4.21 ± 1.48. No significant differences were found in the values of survivin mRNA expression according to histological classification or according to increasing depth of tumor invasion. However, survivin mRNA expression was significantly higher (P 0.01) in patients displaying lymph node metastasis (5.48 ± 1.01) than in patients without the metastasis (4.70 ± 1.55).Conclusions These results indicate that increased survivin mRNA expression begins in the early stages of gastric carcinogenesis. Moreover, the level of survivin mRNA expression may indicate the potential for lymph node metastasis in patients with gastric cancer.     

Hsa-miR-21-3p对食管癌细胞运动与侵袭能力的影响

目的： 研究hsa-miR-21-3p对人食管癌细胞Eca9706侵袭和迁移能力的影响。方法：采用终浓度为30 nmol/L的寡核苷酸hsa-miR-21-3p mimics和阴性对照转染Eca9706细胞48 h后,用实时荧光定量PCR检测转染后hsa-miR-21-3p的表达水平,通过Transwell体外侵袭试验和划痕愈合试验分别观察hsa-miR-21-3p对Eca9706细胞侵袭和迁移能力的影响。结果：与阴性对照组相比较,hsa-miR-21-3p mimics转染组细胞中hsa-miR-21-3p表达明显升高 (P<0.01),是阴性对照的1.1×104倍;Transwell体外侵袭试验结果显示镜下每个视野穿膜细胞数试验组 (76.67±6.11)较阴性对照组 (42.33±2.52)增加了81.1% (P<0.01),表明hsa-miR-21-3p的过表达显著增强食管癌细胞的侵袭能力;划痕愈合试验结果显示,hsa-miR-21-3p试验组的划痕宽度较对照组窄,在相同的时间点,试验组的划痕愈合能力均大于对照组 (P<0.01),说明转染了hsa-miR-21-3p mimics的试验组细胞的迁移能力大于对照组。结论：Hsa-miR-21-3p的高表达可以使人食管癌细胞Eca9706的侵袭和迁移能力增强,提示其可能参与了食管癌进程中的浸润和转移。     

Extra-nodal metastasis should be classified separately from lymph node metastasis in gastric cancer

IntroductionExtra-nodal metastasis (ENM) is defined as a tumor nodule without histological evidence of a lymph node structure. Although ENM has pathological features distinct from those of metastatic lymph nodes, both ENM and metastatic lymph nodes are considered within the same category in the pathological nodal (pN) classification. This study aimed to clarify the clinicopathological characteristics and prognostic relevance of ENM in gastric cancer patients who underwent curative gastrectomy.Materials and methodsWe retrospectively evaluated 1207 Japanese patients who underwent curative gastrectomy at a single center between January 2009 and December 2013. All resected specimens were fixed in 10% formalin, processed, and stained using hematoxylin and eosin, and subsequently reviewed by two pathologists. Survival times were analyzed using the Kaplan-Meier method, and independent prognostic factors were identified using a Cox proportional hazards regression model.ResultsPatients who were ENM-positive had significantly poorer overall survival; multivariable analysis revealed that independent prognostic factors were older age (hazard ratio [HR]: 3.68, 95% confidence interval [CI]: 2.60–5.20), higher pathological tumor classification (HR: 2.28, 95% CI: 1.43–3.62), presence of metastatic lymph nodes (HR: 1.57, 95% CI: 1.0–2.36), and ENM-positive status (HR: 2.33, 95% CI: 1.48–3.66). ENM-positive patients had similar survival outcomes to those of ENM-negative patients with ≥16 metastatic lymph nodes.ConclusionsAmong Japanese patients with gastric cancer who underwent curative gastrectomy, ENM was an independent prognostic factor with a prognostic significance different from that of lymph node metastasis. These results suggest that ENM and lymph node metastasis should be classified separately.     

Protein predictive signatures for lymph node metastasis of gastric cancer

Lymph node status remains one of most crucial indicators of gastric cancer prognosis and treatment planning. Current imaging methods have limited accuracy in predicting lymph node metastasis. We sought to identify protein markers in primary gastric cancer and to define a risk model to predict lymph node metastasis. The Protein Pathway Array (PPA) (initial selection) and Western blot (confirmation) were used to assess the protein expression in a total of 190 freshly frozen gastric cancer samples. The protein expression levels were compared between samples with lymph node metastasis (n = 73) and those without lymph node metastasis (n = 57) using PPA. There were 27 proteins differentially expressed between lymph node positive samples and lymph node negative samples. Five proteins (Factor XIII B, TFIIH p89, ADAM8, COX‐2 and CUL‐1) were identified as independent predictors of lymph node metastasis. Together with vascular/lymphatic invasion status, a risk score model was established to determine the risk of lymph node metastasis for each individual gastric cancer patient. The ability of this model to predict lymph node metastasis was further confirmed in a second cohort of gastric cancer patients (33 with and 27 without lymph node metastasis) using Western blot. This study indicated that some proteins differentially expressed in gastric cancer can be selected as clinically useful biomarkers. The risk score model is useful for determining patients' risk of lymph node metastasis and prognosis.     

非小细胞肺癌组织miRNA-142-3p表达及预后的相关性

目的:探讨miRNA-142-3p在可手术的非小细胞肺癌组织中的表达情况及其与预后的相关性.方法:回顾性分析辽宁省肿瘤医院2009年12月-2012年3月期间行手术治疗的121例非小细胞肺癌患者的临床资料.采用定量RT-PCR检测上述病例肺癌标本中miRNA-142-3p的表达水平,应用 χ2检验分析miRNA-142-3p表达水平与肺癌患者各临床病理参数的相关性,应用Kaplan-Meier方法分析miRNA-142-3p表达与非小细胞肺癌生存的相关性,采用Cox风险比例模型分析预后的独立影响因素.结果:miRNA-142-3p表达量与复发转移情况相关(P<0.05),与患者性别、年龄、病理类型、分期及吸烟史无明显相关,生存分析显示miRNA-142-3p高表达组生存期明显低于低表达组(P<0.01),Cox风险比例模型分析显示miRNA-142-3p表达可作为非小细胞肺癌患者预后的独立影响因素.结论:miRNA-142-3p高表达与非小细胞肺癌复发转移相关,miRNA-142-3p高表达提示非小细胞肺癌患者预后不良.     

Gene expression profiles in human gastric cancer: expression of maspin correlates with lymph node metastasis

To seek for a candidate gene that would regulate tumour progression and metastasis in gastric cancer, we investigated gene expression profiles by using DNA microarray. Tumour tissue and adjacent normal tissue were obtained from 21 patients with gastric cancer and then examined for their gene expression profiles by the Gene Chip Human U95Av2 array, which includes 12 000 human genes and EST sequences. A total of 25 genes were upregulated and two genes were downregulated by at least four-fold in the tumour tissue. In a further analysis according to lymph node metastasis, the expressed levels of maspin, as well as carcinoembryonic antigen and nonspecific crossreacting antigen were significantly higher in tumours with lymph node metastasis than in those without it. Maspin expression in 85 gastric cancer patients was further investigated by using immunohistochemistry. Maspin expression was not observed in normal gastric epithelia without intestinal metaplasia. In contrast, maspin was expressed in 74 of 85 tumour tissues. There was a significant correlation between the incidence of maspin-positive tumour staining and lymph node metastasis. These results suggest that maspin has a potential role for tumour metastasis in gastric cancer.     

Clinicopathological variables associated with lymph node metastasis in submucosal invasive gastric cancer

Background We aimed to elucidate clinicopathological variables associated with lymph node metastasis of submucosal invasive gastric cancer. Methods Specimens were surgically resected from 201 patients who had primary submucosal gastric cancer. We studied 39 consecutive patients with lymph node metastasis and 162 patients without lymph node metastasis. We compared the following clinicopathological characteristics of the patients in relation to lymph node metastasis: age, sex, tumor size, histology, extent of submucosal invasion, lymphatic and venous invasion, and ulceration of the tumor. Submucosal invasion was divided subjectively into sm1, sm2, and sm3 (representing invasion of the upper-, middle-, and lower-third of the submucosa, respectively). We also studied the relationship between lymph node metastasis of submucosal gastric cancer and immunohistochemistry for p53, Ki67, vascular endothelial growth factor (VEGF), α-fetoprotein, sLe^a, and dendritic cells (DCs). Results In terms of conventional pathological factors, lymph node metastasis in submucosal gastric cancer was related to tumor size (P = 0.002), depth of submucosal invasion (P = 0.001), lymphatic invasion (P < 0.0001), and venous invasion (P = 0.012). Lymph node metastasis in sm1 gastric cancer was significantly related to VEGF expression (P = 0.047). Also, lymph node metastasis in sm3 gastric cancer was significantly correlated with DC expression (P = 0.016). Multivariate analysis showed that tumor size, tumor invasion depth in the submucosal layer, and lymphatic invasion were independent predictors of nodal metastasis in submucosal gastric cancer. Conclusion Conventional pathological factors, such as tumor size, depth of submucosal invasion, and lymphatic invasion, have a significant influence on lymph node metastasis. VEGF expression and DC expression may be helpful predictors of lymph node metastasis in patients with sm1 and sm3 gastric cancer, respectively.     

Decrease in ICAM-1 expression on gastric cancer cells is correlated with lymph node metastasis

Background. Lymph node metastasis is a frequent type of metastasis in patients with gastric cancer. The mechanisms responsible for this type of metastasis, however, are not clearly understood. We hypothesize that the immunosurveillance system between cancer cells and lymphocytes may be associated with the lymph node metastatic process. In this study, we examined the correlation between lymph node metastasis and intercellular adhesion molecule-1 (ICAM-1), which mediates the immunosurveillance system between tumor cells and cytotoxic lymphocytes, in gastric cancer. Methods. One hundred and forty-three specimens resected from patients with gastric cancer were investigated by staining with a monoclonal antibody against ICAM-1. We studied the correlation between the expression of ICAM-1 and various clinicopathologic factors, as well as infiltration of tumor-infiltrating lymphocytes (TILs). Results. ICAM-1 expression on gastric cancer cells was significantly decreased in patients with lymph node metastasis. The infiltration of TILs was associated with ICAM-1 expression level. The prognosis of patients with ICAM-1-negative tumors was poorer than the prognosis of those with ICAM-1-positive tumors. Conclusions. These findings suggest that ICAM-1 expression on cancer cells is closely associated with lymph node metastasis in gastric cancer, under the influence of the host immunosurveillance system. Received on Aug. 20, 1999; accepted on Jan. 5, 2000     

乳腺癌肿瘤代谢异质性与p53基因表达和淋巴结转移相关性的研究

目的:分析乳腺癌肿瘤代谢异质性与p53基因表达和乳腺癌淋巴结转移的关系。方法:选取接受18F-FDG PET/CT检查的原发性乳腺癌患者34例,分别测定每例患者原发灶的肿瘤代谢异质因子（heterogeneity factor,HF）,同时将HF与p53基因表达和淋巴结转移进行相关性分析,采用受试者工作特性曲线（area under receiver operating characteristic curve,ROC）分析HF对p53基因突变及淋巴结转移的诊断效能。结果:HF与乳腺癌p53基因表达呈弱相关性（P<0.05）;HF与淋巴结转移无相关性（P>0.05)。结论:HF与乳腺癌p53基因表达具有相关性,HF对预测p53基因表达具有一定意义,然而,HF不能预测乳腺癌是否有淋巴结转移。